Figure 4

Optimal culturing conditions for highly efficient transduction of primary CD45RA⁺T cells. A. Virus was generated using the standard production and concentration protocol. Primary CD45RA⁺ T cells were stimulated using plate bound anti-CD3 (5 μg/mL) and soluble anti-CD28 (1 μg/mL) or with CD3/CD28 activation beads (1 bead to 5 cells) then transduced following 24 hrs of stimulation. ***P < 0.001 as determined by Students t-test. Data represent mean ± s.d. of three independent experiments. B. Primary CD45RA⁺ T cells were stimulated with CD3/CD28 activation beads for 24 hrs then transduced and cultured in X-VIVO media supplemented with 10% FCS or 10% HS. ***P < 0.001 as determined by students t-test. Data represent mean ± s.d. of three independent experiments. C and D. Primary CD45RA⁺ T cells were stimulated using CD3/CD28 activation beads and cultured in X-VIVO media supplemented with 10% HS and transduced following 0, 24 and 48 hrs of activation. *P < 0.05 as determined by one-way ANOVA with Bonferroni correction. Data represent mean ± s.d. of three independent experiments.